Poison ivy hairy root cultures enable a stable transformation system suitable for detailed investigation of urushiol metabolism

dc.contributor.authorLott, Aneirin A.en
dc.contributor.authorFreed, Catherine P.en
dc.contributor.authorDickinson, Christopher C.en
dc.contributor.authorWhitehead, Susan R.en
dc.contributor.authorCollakova, Evaen
dc.contributor.authorJelesko, John G.en
dc.date.accessioned2020-09-11T19:30:39Zen
dc.date.available2020-09-11T19:30:39Zen
dc.date.issued2020en
dc.description.abstractPoison ivy (Toxicodendron radicans) is best known for causing exasperating allergenic delayed-contact dermatitis symptoms that last for weeks on persons who have contacted the plant. Urushiols are alkylcatechols produced by poison ivy responsible for causing this dermatitis. While urushiol chemical structures are well known, the metabolic intermediates and genes responsible for their biosynthesis have not been experimentally validated. A molecular genetic characterization of urushiol biosynthesis in poison ivy will require stable genetic transformation and subsequent regeneration of organs that retain the capacity synthesize urushiol. To this end, Agrobacterium rhizogenes was used to generate hormone-independent poison ivy hairy root cultures. Optimal conditions for hairy root formation were skotomorphic poison ivy hypocotyls prick-inoculated with A. rhizogenes, and preferential propagation of cultures with an atypical clumpy hairy root growth habit. The origin of the poison ivy accession used for A. rhizogenes prick-inoculation did not affect the initial formation of calli/ hairy root primordia, but rather significantly influenced the establishment of longterm hormone-independent hairy root growth. A. rhizogenes harboring a recombinant T-DNA binary plasmid with an intron-containing Firefly Luciferase gene produced stable transgenic hairy root lines expressing luciferase activity at high frequency. Poison ivy hairy root lines produced significantly lower steady-state urushiol levels relative to wild-type roots, but higher urushiol levels than a poison ivy undifferentiated callus line with undetectable urushiol levels, suggesting that urushiol biosynthesis requires intact poison ivy organs. The lower urushiol levels in poison ivy hairy root lines facilitated the first identification of anacardic acid metabolites initially in hairy roots, and subsequently in wild-type roots as well. This study establishes a transformation hairy root regeneration protocol for poison ivy that can serve as a platform for future reverse-genetic studies of urushiol biosynthesis in poison ivy hairy roots.en
dc.description.sponsorshipVirginia Tech Global Change Center Undergraduate Awarden
dc.description.sponsorshipVirginia Tech Translational Plant Interdisciplinary Graduate Education Programen
dc.description.sponsorshipirginia Tech School of Plant and Environmental Sciencesen
dc.description.sponsorshipVirginia Tech Foundation LLCen
dc.description.sponsorshipVirginia Tech Biological Sciences Department Start-Up Fundsen
dc.format.mimetypeapplication/pdfen
dc.identifier.doihttps://doi.org/10.1002/pld3.243en
dc.identifier.urihttp://hdl.handle.net/10919/99946en
dc.language.isoenen
dc.publisherWileyen
dc.rightsCreative Commons Attribution-NonCommercial-NoDerivatives 4.0 Internationalen
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/4.0/en
dc.subjectAgrobacterium rhizogenesen
dc.subjectanacardic aciden
dc.subjecthairy rootsen
dc.subjectToxicodendron radicansen
dc.subjecttransformationen
dc.subjecturushiolen
dc.titlePoison ivy hairy root cultures enable a stable transformation system suitable for detailed investigation of urushiol metabolismen
dc.title.serialPlant Directen
dc.typeArticle - Refereeden
dc.type.dcmitypeTexten

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